We have been studying the K562 human leukemia cell line in order to analyze the control of human erythroid differentiation and globin gene expression. We have shown that hemin causes induction of hemoglobin synthesis by increasing mRNA levels and that this process is fully reversible, without evidence of terminal differentiation. We have studied the physiological properties of the embryonic and fetal hemoglobins synthesized in these cells as a response to hemin. We find that following induction of hemoglobin synthesis K562 cells demonstrate oxygen dissociation properties similar to human embryonic red cells. The P50 value of the hemoglobin is 20 plus and minus 0.9 mmHg. The Hill coefficient is 2.5. Intracellular pH of K562 cells, reasured by 3l-P NMR, is 7.3. We also find that the levels of 2,3-diphosphoglycerate (DPG), a potent modifier of hemoglobin function, increase dramatically upon the induction of hemoglobin synthesis in K562 cells. In a typical experiment, DPG levels went from 8 nmoles/one hundred million cells to 64 nmoles/one hundred million cells upon induction of hemoglobin accumulation. For several experiments there was an approximately linear relationship between DPG and hemoglobin levels. The coordinate increase in DPG and hemoglobin levels has been demonstrated not only by an enzymatic method but also by 3l-P NMR spectroscopy. These data suggest that DPG and hemoglobin levels are coordinately controlled in these cells. We are now investigating the molecular mechanism of the coordinate increase in DPG and hemoglobin levels in response to hemin induction. In more recent experiments we have examined the effects of various analogs of 5-azacytidine which may also inhibit DNA methylation for their ability to induce hemoglobin synthesis in K562 cells. We find that 3-deazadenosine and several other analogs do cause induction. We are examining the molecular basis of this phenomenon as it may be relevant to the treatment of sickle cell anemia and thalassemia patients in an effort to increase fetal hemoglobin synthesis.